Mucosal Immunity

Abstract Despite advances in our understanding of immunoglobulin A nephropathy (IgAN) over the past decade, there are currently no specific therapies capable of targeting key pathways involved in the pathogenesis of the disease. Recent studies have, however, provided new insights into important molecular pathways that are likely to be amenable to therapeutic manipulation in the future. Specifically, a deeper understanding of the role of mucosal immunity, B‐cell activation and mesangial cell activation in IgAN has provided the impetus for a number of exciting phase II/III clinical trials in IgAN. In this review, we examine some of these on‐going studies, first examining studies that clarify the role of traditional immunosuppression in IgAN, then focusing on novel therapies in early clinical studies, looking closely at the rationale for these agents in relation to our current understanding of the pathogenesis of IgAN. Finally, we examine emerging pathways and therapeutic agents that have the potential to be developed as novel therapies in the coming years. It is hoped that as we continue to develop a greater understanding of IgAN, emerging therapies will soon become a reality in the day‐to‐day treatment of patients with IgAN. Immunoglobulin A nephropathy (IgAN) is the commonest pattern of glomerulonephritis in the world and an important cause of end‐stage renal disease (ESRD).1, 2 Since its first description more than 47 years ago,3 there have been significant advances in our understanding of the pathogenesis of IgAN, with numerous studies offering key insights into the molecular mechanisms of the disease.4 There is now convincing evidence that the production of poorly galactosylated IgA1 and glycan‐specific IgG and IgA autoantibodies leads to the formation of IgA‐containing immune complexes, and these immune complexes deposit within the mesangium, where their consequent effects on mesangial cells, podocytes, and proximal tubule cells are central to the development of IgAN. Despite gaining better insight into the pathogenesis of IgAN, current treatment strategies remain generic and common to all forms of chronic kidney disease, principally renin–angiotensin system (RAS) blockade, reduction of proteinuria and blood pressure control, or involve the use of non‐specific immunosuppression, such as corticosteroids.5 According to the recently published KDIGO guidelines (2012), RAS blockade with angiotensin‐converting enzyme inhibitors (ACEi) or angiotensinogen receptor blockers (ARB) when proteinuria is more than 1 g/day remains the mainstay of treatment in IgAN. Theoretically, more intense and complete RAS inhibition using more than one RAS blocker versus a single agent could offer more benefit in patients with proteinuric renal disease. However, following the retraction of the COOPERATE study, there are no studies looking specifically at dual RAS blockade in patients with IgAN.6 In addition, there is emerging evidence that dual blockade, in specific groups of patients, with kidney disease, does not provide additional benefit but actually increases the risk of adverse events.7-9 It has been suggested that it is premature to abandon combination RAS blockade on the basis of the current level of evidence and that an approach of individualizing therapy with careful dose titration according to proteinuria and tolerability in patients with kidney disease should be adopted,10 a practice that will require formal testing. Immunosuppression, in the form of a 6 month course of corticosteroid therapy, is only suggested in patients who have persistent proteinuria and preserved renal function (glomerular filtration rate (GFR) >50 mL/min per 1.73 m2) after a 3‐ to 6 month period of optimization of blood pressure control and RAS blockade (grade 2C evidence). At the time the KDIGO guidelines were written, there was insufficient evidence to support the use of any other form of immunosuppression (apart from in crescentic IgAN). With the exception of the recommendation on the use of RAS blockade when proteinuria exceeds 1 g/day, the remaining KDIGO suggestions for the treatment of IgAN were based on low levels of evidence (grade 2B–2D). The lack of high‐quality clinical trials has limited the robust evaluation of therapies in IgAN, particularly immunosuppressive regimens, and there remains a great deal of uncertainty over the optimum treatment of patients at high risk of progressive chronic kidney disease. In addition, current therapeutic options seek to ameliorate the damage occurring as a consequence of inflammation and scarring far downstream in the disease pathology, while disease‐specific therapies capable of targeting early events in the pathogenesis of IgAN, for example, preventing IgA immune‐complex formation or mesangial IgA deposition and mesangial cell activation, are currently lacking. In this context, there is a pressing need for a better understanding of the role of immune modulation in IgAN and in evaluation of novel treatment strategies that target specific pathogenic pathways beyond current traditional treatments. One of the challenges in the evaluation of treatment options is, like many other kidney diseases, IgAN is frequently slowly progressive, and high‐risk patients may develop ESRD only after many years. Significant loss of renal function (defined by the doubling of serum creatinine) has been accepted as a valid end point for clinical studies, but even this is a relatively long‐term outcome. Surrogate end points, such as proteinuria, thus become an important tool in the evaluation of clinical studies as these allow more rapid prediction of clinically important outcomes using smaller numbers of patients over a shorter period of time.11 In IgAN, proteinuria has been demonstrated to be an early marker of progressive disease, and treatment resulting in reduction of proteinuria predicts a favourable long‐term outcome.12 However, there is uncertainty over the use of proteinuria as a surrogate end point in clinical studies. First, while it has been demonstrated that proteinuria is a marker of kidney damage and predicts risk of progression in renal disease, it remains unclear if proteinuria causes or mediates progression of kidney disease. Furthermore, proteinuria is not always a necessary intermediate endpoint on the path to ESRD, implying that its relationship to progressive renal disease is not always direct. Intervention trials demonstrating association between effects of treatment on proteinuria and renal outcome are either limited to specific agents (e.g. ACEi) or specific diseases (e.g. diabetic nephropathy), and the results cannot necessarily be extrapolated to other kidney diseases or pharmacologic agents. This is especially so when considering that kidney disease is heterogeneous and the degree of proteinuria (and its consequent effects) varies widely. Finally, the size effect of proteinuria as a surrogate marker in predicting renal outcomes is not precisely defined, meaning clinicians cannot quantify reliably the effects on renal outcome based on the degree of reduction in proteinuria. For these reasons, regulatory agencies, such as the FDA, have been cautious when reviewing approvals based predominantly on a surrogate end point of proteinuria as evidence of treatment efficacy for drug approvals.13 Therefore, in the absence of a sensitive and specific mechanistic biomarker that mirrors disease activity or predicts response to treatment, the definition of the primary end point in many current clinical trials in IgAN remains contentious, balancing the pragmatic need to conduct a meaningful trial while appeasing regulators to allow early licensing of the drug under investigation. As you will see from Tables 1-7, most investigators have opted to use proteinuria reduction as a surrogate marker of reduced risk of renal function decline and ESRD. It remains to be seen how regulatory agencies will view relatively short‐term reductions in proteinuria as evidence of efficacy of emerging therapies in IgAN. Inclusion criteria All biopsy‐proven IgAN patients, between 18 and 70 years of age will be considered if: Proteinuria >0.75 g/day, and Hypertension (BP > 140/90 mm Hg or use of antihypertensive medications), or GFR < 90 mL/min Exclusion criteria GFR < 30 mL/min At the end of run‐in phase, patients will be excluded if: Proteinuria >3.5 g/day, or Decrease in GFR > 30% (from baseline) Trial design Intervention Follow‐up duration Run‐in phase All patients will receive supportive therapy: RAS blockade, BP control (target 125/75 mm Hg), statin therapy, dietary counselling for low salt diet and protein restriction (with verification of dietary restriction via measurement of 24‐hour urine sodium and urea excretion) and education/intervention programme to stop smoking 6 months Randomization phase Patients with persistent proteinuria exceeding 0.75 g/day at the end of the run‐in phase will be randomized to two groups: Supportive group Supportive group will continue supportive therapy as initiated in the run‐in phase Immunosuppressive group GFR ≥ 60 mL/min Six month course of corticosteroids, consisting of intravenous methylprednisolone 1 g per day for 3 days at the start of month 1, 3, 5 and oral prednisolone 0.5 mg/kg every other day for 6 months on the remaining days GFR between 30 and 60 mL/min Oral cyclophosphamide (1.5 mg/kg per day, adjusted down to the nearest 50 mg) for 3 months, together with oral prednisolone (40 mg/day, tapered to 10 mg at 3 months), after which patients will receive azathioprine (1.5 mg/kg per day), together with oral prednisolone (10 mg/day for months 4–6 and 7.5 mg/day after 6 months) for 3 years 3 years Primary end point Complete remission defined by proteinuria of less than 0.2 g/day with stable renal function (loss of GFR less than 5 mL/min from baseline) GFR loss of 15 mL/min or higher, at the end of 3 years BP, blood pressure; GFR, glomerular filtration rate; IgAN, IgA nephropathy. Inclusion criteria All biopsy‐proven IgAN patients, above 14 years of age will be considered if: Proteinuria >1 g/day, and GFR 20–90 mL/min Exclusion criteria Received immunosuppression in past 1 year Trial design Intervention Follow‐up duration Pre‐randomization phase Dose of ACEi or ARB and blood pressure will be optimized 4–8 weeks Randomization phase Patients with persistent proteinuria exceeding 1 g/day at the end of the pre‐randomization phase will be randomized to two groups: Oral methylprednisolone 0.8 mg/kg per day for 2 months (rounded to the nearest 4 mg and with a maximal dose of 48 mg/day) then tapered by 8 mg/day each month Matching placebo at the same dosage 6–8 months Primary end point Composite of: 50% decrease in GFR Development of ESRD, and Death due to kidney disease. The planned average follow‐up period is for a duration of 5 years, although the study is event driven and will be continued until 335 primary endpoints have occurred, so the final follow‐up duration may be longer or shorter depending on the event rate. ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; BP, blood pressure; ESRD, end‐stage renal disease; GFR, glomerular filtration rate; IgAN, IgA nephropathy. Inclusion criteria All biopsy‐proven IgAN patients, above 18 years of age will be considered if: UPCR > 0.5 g/g or 24‐UTP > 0.75 g/day, and GFR > 45 mL/min Exclusion criteria Received immunosuppression in past 2 years At the end of run‐in phase, patients will be excluded if: Decrease in GFR > 30% (from baseline) Trial design Intervention Follow‐up duration Run phase ACEi and/or ARB will be dosed to target a blood pressure of 130/80 mm Hg and UPCR < 0.5 g/g 6 months Randomization phase Patients with persistent proteinuria exceeding UPCR > 0.5 g/g at the end of the run‐in phase will be randomized to three groups: Enteric budesonide 16 mg/day, Enteric budesonide 8 mg/day, or Matching placebo group Patients who received 16 mg/day dosing will taper the dose to 8 mg/day during the first 2 weeks of the follow on phase, with the placebo and 8 mg/day groups receiving placebo to maintain blinding. 9‐month treatment and 3‐month follow‐on Primary end point Change in UPCR at 9 months ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; BP, blood pressure; GFR, glomerular filtration rate; IgAN, IgA nephropathy; UPCR, urine protein : creatinine ratio; UTP, urine total protein. Inclusion criteria All biopsy‐proven IgAN patients, between 18 and 65 years of age will be considered if: Proteinuria between 1 and 6 g/day GFR > 30 mL/min Stable dose of ACEi and/or ARB prior to screening Exclusion criteria Received immunosuppression within past 6 months Trial Design Intervention Follow‐up duration Randomization phase Patients will be randomized to two groups: Treatment group: Induction phase – subcutaneous blisibimod 100 mg three times weekly for 8 weeks, then Maintenance phase – subcutaneous blisibimod 200 mg weekly for 16 weeks Placebo group 24 weeks Primary end point Change in proteinuria at 24 weeks ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; BP, blood pressure; GFR, glomerular filtration rate; IgAN, IgA nephropathy. Inclusion criteria All biopsy‐proven IgAN patients, between 18 and 70 years of age will be considered if: Proteinuria >0.5 g/day GFR > 30 mL/min Stable dose of ACEi and/or ARB for at least 90 days prior to randomization Exclusion criteria Prior use of immunosuppressant Trial design Intervention Follow‐up duration Randomization phase Patients will be randomized to three groups: Fostamatinib 150 mg twice daily, Fostamatinib 100 mg twice daily, or Placebo group 24 weeks Primary end point Change in proteinuria at 24 weeks ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; GFR, glomerular filtration rate; IgAN, IgA nephropathy. Inclusion criteria All biopsy‐proven IgAN patients, between 18 and 70 years of age will be considered if: Proteinuria >1 g/day (or >0.5 g/day if on combination RAS blockers) GFR 30 to 90 mL/min Exclusion criteria Received >6 months of corticosteroids Trial design Intervention Follow‐up duration Randomization phase Patients will be randomized to two groups: Intravenous rituximab 1 g on day 1 and 15. The course will be repeated again at 6 months (i.e. day 168 and 182) after the first infusion Placebo All patients will receive, in addition to ACEi and/or ARBs, omega‐3 fatty acid fish oil supplements 3.6 g EPA/day 12 months Primary end point Change in proteinuria at 12 months Patients will be analysed according to response – complete response defined as proteinuria <300 mg/day and less than 10% decline in GFR from baseline, and partial response defined as reduction of proteinuria >50% from baseline and decline in GFR of less than 25% from baseline Patients with proteinuria reduction of less than 50%, unchanged or increasing proteinuria and/or decline in renal function will be classified as non‐responder. ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; EPA, eicosapentaenoic acid; GFR, glomerular filtration rate; IgAN, IgA nephropathy; RAS, renin–angiotensin system. Inclusion criteria All biopsy‐proven IgAN patients, above 18 years of age, will be considered if: Proteinuria >1 g/day Stable dose of ACEi and/or ARB for at least 4 weeks prior to screening Trial design Intervention Follow‐up duration Open‐label study Patients will receive intravenous bortezomib 1.3 mg/m2 on days 1, 4, 8 and 11 A second cycle is to be given a month later for non‐responders 1 year Primary end point Reduction in proteinuria at 1 year ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; IgAN, IgA nephropathy. In this article, we review the on‐going clinical trials in IgAN that examine both traditional treatments and emerging therapies, looking closely at the rationale, design and the progress of individual studies, and finally, we will examine some of the potential new therapies that hold promise but are not yet in clinical trials in IgAN. We have summarized our current understanding of the pathogenesis of IgAN in Figures 1 and 2 and have included the sites of action of the emerging therapies in relation to our understanding of key pathogenic pathways in IgAN. STOP‐IgAN (ClinicalTrials.gov Identifier: NCT00554502) Rationale The formation of IgA‐containing immune complexes is central to the pathogenesis of IgAN and suggests that modulation of B and T cell function with immunosuppressants may be beneficial in IgAN. A number of small clinical trials have assessed the value of immunosuppression (mostly involving corticosteroids) in IgAN; however, a major limitation of these older studies is the lack of conformity in the use of RAS inhibition and blood pressure targets with current best practice. This is likely to have led to the inclusion of patients who would have achieved a reduction in proteinuria and stabilization of GFR with RAS inhibition and blood pressure control alone. It is currently unclear, therefore, whether conventional immunosuppressive therapy adds benefit above current best practice supportive care. The STOP‐IgAN (Supportive Versus Immunosuppressive Therapy of Progressive IgA Nephropathy) trial was designed to specifically evaluate the additive benefit of traditional immunosuppressive therapy to current best practice standard care.14 Trial design, inclusion and exclusion criteria, and primary end point The STOP‐IgAN trial is a phase III prospective, randomized, multi‐centre, open‐label study conducted in Germany, comparing two parallel arms: immunosuppressive therapy added to comprehensive supportive therapy or comprehensive supportive therapy alone. Briefly, the trial encompasses a 6 month run‐in phase where patients receive best supportive therapy. Patients with persistent proteinuria exceeding 0.75 g/day at the end of the run‐in phase are randomized to either continue supportive therapy alone or to receive supportive therapy and immunosuppression. Patients randomized to the immunosuppression arm receive one of two‐treatment regimens, depending on their renal function (Table 1). Trial progress to date The trial began in 2008, and recruitment was completed in 2014 (148 patients – 74 in each arm). Interestingly, only slightly more than one‐third of the patients screened were eligible for randomization as there was a significant reduction of proteinuria during the run‐in phase with supportive treatment alone. This observation reinforces the fact that many IgAN patients with significant proteinuria can achieve a significant reduction in proteinuria (<0.75 g/d) with optimal supportive care only. Preliminary results from the study was announced at the 52nd ERA‐EDTA congress (Late Breaking Clinical Trials session) in London. The number of patients achieving complete remission was higher in the group receiving immunosuppression (odds ratio 5.33, P = 0.008, based on available case analysis); however, this was not accompanied by a difference in GFR decline between the two groups at three years (−1.6 versus −1.5 ml/min in supportive and immunosuppression group respectively, P = 0.362). Furthermore, treatment with immunosuppression was associated with higher rate of serious adverse events such as infections, diabetes and weight gain. The full report of the study is expected to be published soon. TESTING (ClinicalTrials.gov Identifier: NCT01560052) Rationale To date, there have been several small studies specifically investigating the use of corticosteroids in IgAN. Of these studies, three found that corticosteroids were associated with reduced progression of IgAN. In the largest published randomized controlled trial by Pozzi et al., treatment with corticosteroids was associated with a reduction in proteinuria and reduction in the risk of ESRD over a 10 year period.15 However, RAS blockade was used only in a minority of the patients, and blood pressure control was sub‐optimal by current standards. In two more recent trials, treatment with oral prednisolone for 6–8 months (a regimen less intense than that used by Pozzi et al.) resulted in a reduction in the proportion of patients progressing to doubling of serum creatinine or ESRD.16, 17 However, in both trials, patients stopped RAS blockade prior to entering the trial, and there was no run‐in phase before randomization to optimize the ACEi and/or ARB dose. It is therefore possible that the patients in these trials might have responded to ACEi/ARB alone (a hypothesis being tested by the STOP‐IgAN trial detailed above). In a recent systematic review, Lv et al. identified nine trials examining the effect of steroids in IgAN (536 IgAN patients with proteinuria >1 g/day and normal renal function at inclusion). The use of steroid therapy was associated with a lower risk of doubling of serum creatinine/halving of GFR or ESRD. In these studies, steroid therapy was, however, associated with a 55% higher risk of adverse events, including diabetes, weight gain and other features of steroid excess. Many of the included studies were single‐centre, comprised small numbers of patients and had only short‐term follow‐up data. Therefore, the balance of benefits and risks of corticosteroid treatment in IgAN remains uncertain, and it is not clear who should be treated with this therapy. The Therapeutic Evaluation of STeroids in IgA Nephropathy Global (TESTING) trial was designed to specifically answer this question. Trial design, inclusion and exclusion criteria and primary end point The TESTING study is an international, multi‐centre, double‐blind, randomized, placebo‐controlled trial to study the role of oral methylprednisolone in preventing kidney events in high‐risk IgAN patients. Unlike previous studies, this study includes patients with a GFR between 20 and 90 mL/min per 1.73 m2, and as with STOP‐IgAN, patients enrolled in the TESTING study enter a pre‐randomization period where the dose of ACEi or ARB and blood pressure is optimized over a 4‐ to 12 week period (depending on whether the patient was on stable dose of ACEi or ARB prior to the pre‐randomization period). Table 2 summarizes the design of the study. The target sample size for the study is 1300 patients (650 in each group) and will make this study the largest randomized controlled clinical trial ever performed in IgAN. Trial progress to date The study began recruitment in 2012, and randomization is expected to be complete in 2017. NEFIGAN (ClinicalTrials.gov Identifier: NCT01738035) Rationale One of the key observations in patients with IgAN is the presence of poorly galactosylated IgA1 in both the serum and glomerular immune deposits, a finding that is consistently reproduced in populations of different geographic origin. The precise origin of this IgA1 remains the subject of ongoing investigations.18-21 A number of observations have, however, led to the postulation that the mucosal immune system may be an important source of poorly galactosylated IgA1. It has been shown that the site of antigen encounter influences the phenotype of B lymphocytes and the IgA that they subsequently produce. In health, B cells that encounter antigen in the systemic compartment (e.g. following subcutaneous or intradermal immunization) synthesize heavily galactosylated IgA1 while mucosally programmed B cells, which encounter antigen across mucosal surfaces, synthesize poorly galactosylated IgA1.22 It is therefore postulated that the poorly galactosylated IgA1 in IgAN may originate from the mucosal immune system as an exaggerated mucosally primed B cell response to commonly encountered mucosal pathogens. Whether this response is limited to the mucosa‐associated lymphoid tissue or involves B cells that have mis‐trafficked to the bone marrow is currently not known.23, 24 A number of separate studies have reported high levels of IgA antibodies in the serum against a variety of mucosal antigens including Helicobacter pylori, oral polio vaccine, gliadin, ovalbumin and dextran B512 in patients with IgAN.25-29 Furthermore, the well‐recognized association of mucosal infection with episodes of visible haematuria suggests a close relationship between the mucosal immune system and nephritogenic IgA1 production in IgAN. Consistent with these observations, recently published genome‐wide association studies in IgAN have identified loci that are directly associated with intestinal mucosal immunity.30, 31 Therefore, given that the mucosal immune system may play a key role in the generation of nephritogenic IgA in IgAN, immunosuppression targeted to sites of mucosal B‐cell induction may provide an alternative to traditional regimens of systemic immunosuppression which are associated with significant side effects. In an open‐label, uncontrolled, proof‐of‐concept pilot study by Smerud et al., the potential treatment effects and safety profile of Nefecon® – a modified release formulation of budesonide, specifically designed to deliver budesonide to the ileocecal Peyer's patches, with minimal systemic exposure and side effects – were examined.32 Sixteen patients (62.5% males) with biopsy‐proven IgAN (albuminuria >500 mg/day and serum creatinine <200 μmol/L) were treated with Nefecon® 8 mg/day for 6 months and followed up for a further 3 months. During the 6 month treatment period, there was a median relative reduction in urinary albumin excretion of 23% (interquartile range −0.36 to −0.4, P = 0.04). This was accompanied by a minor reduction (6%) of serum creatinine and modest increase (8%) of GFR. Three out of 16 patients were withdrawn prematurely from the study due to abdominal pain relating to the study drug (n = 2) and sleep disturbances and increased micturition (n = 1). No major corticosteroid‐related side effects were reported. This study, coupled with our current understanding of the role of mucosal immunity in IgAN, formed the basis for the NEFIGAN trial. Trial design, inclusion and exclusion criteria, and primary end point The NEFIGAN trial is a multi‐centre, double‐blind, randomized, placebo‐controlled phase 2b study to evaluate the efficacy and safety of two different doses of Nefecon® in the treatment of patients with primary IgAN. The study is being conducted in 62 centres in 10 European countries. Consistent with other studies, there is an initial phase of rigorous blood pressure control and maximization of RAS blockade over a 6 month run‐in phase. Patients are eligible for randomization into the treatment phase of the trial (Table 3) if they have a UPCR of >0.5 g/g despite maximal supportive care. Trial progress to date The study began in 2012, and recruitment of patients was completed in January 2014. In contrast to the STOP‐IGAN trial, of the 299 patients who entered screening, 70% (208) were eligible for randomization suggesting that in the 4 years from commencement of STOP‐IgAN to 2012, there had been a significant improvement in the standard of care received by patients with IgAN across Europe. Results from this study are expected in mid‐2015. On the 14 April 2015, Pharmalink announced that the NEFIGAN trial had fully met its primary efficacy end point at a planned interim analysis and had been stopped early with respect to statistical analysis of the endpoint. BRIGHT‐SC (ClinicalTrials.gov Identifier: NCT02062684) Rationale Effective B cell maturation and survival is dependent on the presence of BAFF (B cell activating factor), also known as B‐lymphocyte stimulator (BlyS), and APRIL (a proliferation inducing ligand). BAFF and APRIL are expressed by T cells, dendritic cells, monocytes and macrophages but not by B cells. BAFF and APRIL bind to the TNF superfamily receptors B cell maturation antigen, TNFR homolog transmembrane activator and Ca2+ modulator and CAML interactor (TACI) and B‐cell‐activating factor receptor (does not bind APRIL). These receptors are expressed almost exclusively by immune cells in the B cell lineage. Expression is weak on immature bone marrow B cells but becomes progressively stronger during peripheral B cell development. Ligation of these receptors contributes to T‐cell‐independent responses of B cells, negative regulation of the size of the B‐cell compartment, class‐switch recombination and plasma cell survival.33-35 Serum levels of BAFF are elevated in patients with autoimmune disease, and levels of BAFF correlate with autoantibody levels.36-40 It has also been reported that elevated levels of BAFF are associated with an increased risk of antibody‐mediated renal allograft rejection.41 Evidence is also emerging for a role of BAFF in IgAN. Transgenic mice overexpressing BAFF display B‐cell hyperplasia, have elevated levels of IgA in the serum and intestinal lamina propia, and develop mesangial IgA deposition. Interestingly, mesangial IgA deposition only developed in those mice exposed to environmental antigens, presumably reflecting mucosal B cell activation.42, 43 In a separate study of tonsillar mononuclear cells (TMCs) exposed to deoxycytidyl‐deoxyguanosineoligodeoxynucleotides, to mimic the immunostimulatory activity of microbial DNA, Goto et al. demonstrated that high levels of BAFF were associated with an increased production of IgA by TMC in IgAN.44 Furthermore, the production of IgA by TMCs was inhibited by blockade of BAFF signalling. Serum levels of BAFF are elevated in IgAN, and BAFF levels correlate with a worse outcome in IgAN as measured by renal histology (increased mesangial hypercellularity, segmental glomerulosclerosis and tubular atrophy/interstitial fibrosis) and higher serum creatinine at time of renal biopsy.45 Interruption of BAFF and APRIL signalling with belimumab (Benlysta®, a human monoclonal antibody that inhibits BAFF), blisibimod (a selective peptibody antagonist of BAFF) and atacicept (a fusion protein containing the extracellular, ligand‐binding portion of TACI and the modified Fc portion of human IgG, blocks BAFF and APRIL) has proven effective in phase II/III trials in systemic lupus erythematosus (SLE).46 Given the success of treatment with BAFF inhibitors for patients with SLE,47, 48 there is considerable interest in examining the efficacy of BAFF inhibition in other B cell‐driven diseases, including IgAN. The first such study in IgAN is the BRIGHT‐SC study (Blisibimod Response in IgA Nephropathy Following At‐Home Treatment by Subcutaneous Administration). Trial design, inclusion and exclusion criteria, and primary end point BRIGHT‐SC is an international multi‐centre, randomized, double‐blind, placebo‐controlled phase II/III trial to examine the efficacy and safety of blisibimod in patients with IgAN (Table 4). Blisibimod (a fusion between the Fc portion of IgG and a peptide sequence selected for its ability to bind with high affinity to BAFF) is able to bind both soluble and membrane bound BAFF. Target recruitment for BRIGHT‐SC is 200 patients. Trial progress to date The study began in 2013 in Asia and is currently recruiting patients in Asia and Europe. Recruitment is expected to be complete by 2016. SIGN (ClinicalTrials.gov Identifier: NCT02112838) Rationale Effective B cell maturation and survival are not only dependent on BAFF and APRIL but also signalling through the B cell receptor (BCR). Activation through the BCR is required for normal antibody production, and defects in BCR signal transduction may lead to immunodeficiency, autoimmunity and B‐cell malignancy. One of the key intracellular signal transduction pathways activated on ligation of the BCR is the spleen tyrosine kinase (Syk)–Bruton's tyrosine kinase (BTK) axis, where BTK acts as an essential downstream effector of Syk in regulating both the maturation and survival of the B‐cell lineage. Given the central role of Syk in transmission of activating signals within B‐cells, it has been proposed that Syk plays a key role in autoantibody production and the pathogenesis of autoimmune disorders. In the only published clinical trial to date, administration of fostamatinib, an orally active relatively selective small molecule Syk inhibitor, resulted in a dose‐dependent improvement in disease severity in a 6 month double‐blind, placebo‐controlled trial in rheumatoid arthritis. While Syk is primarily expressed in haematopoietic tissues, there is expression of Syk in a variety of other tissues where it mediates diverse biological functions including cellular adhesion, innate immune recognition, osteoclast maturation, platelet activation and vascular development. This wide tissue distribution may explain the side effects seen with fostamatanib in the rheumatoid arthritis trial. Side effects included diarrhoea, upper respiratory tract infections, neutropenia and hypertension. Interestingly, Syk expression has also recently been reported in the kidney where it appears to regulate the kidney's response to injury.49 In an animal model of nephrotoxic nephritis Syk inhibition reduced the severity of the inflammatory cell infiltrate and proinflammatory cytokine levels in the kidney, reduced the extent of histological damage and reduced the level of proteinuria, even when treatment was delayed until disease was well established.50 In vitro experiments by the same authors demonstrated a down‐regulation of MCP‐1 production when mesangial cells and macrophages were stimulated with aggregated IgG in the presence of Syk inhibitors. Separately, Syk inhibition was shown to significantly reduce autoantibody production in a rodent model of experimental autoimmune glomerulonephritis.51 Similar to the nephrotoxic nephritis model, Syk inhibition decreased the production of proinflammatory cytokines in ex vivo nephritic glomeruli and in vitro bone marrow‐derived macrophages suggesting a therapeutic effect of Syk inhibition independent of its effect on autoantibody production. There is also now evidence that Syk plays a role in determining the mesangial cell response to IgA in IgAN. Kim et al. demonstrated that inhibition of Syk in mesangial cells was capable of blocking the proliferative and pro‐inflammatory effects of IgA immune complexes in IgAN.52 Furthermore, when renal biopsies were stained for total and phosphorylated Syk, there was a clear upregulation of Syk in glomeruli of patients with IgAN. As a result of these experiments and the crucial role of Syk in BCR signalling, the SIGN (Syk Inhibition for Glomerulonephritis) study was developed to evaluate the efficacy of Fostamatinib disodium, the orally bioavailable prodrug of the active compound R406, in the treatment of IgAN.53 Trial design, inclusion and exclusion criteria, and primary end point SIGN is an international, multi‐centre, phase 2, double‐blind, placebo‐controlled, randomized trial on the safety and efficacy of fostamatinib in the treatment of IgAN (Table 6). The study aims to recruit 75 patients, and notably, patients will undergo a repeat renal biopsy after treatment to evaluate the effects of Syk inhibition on histopathology. Trial progress to date The first patient was randomized in late 2014, and recruitment is expected to be complete by 2016. Rituximab in IgAN (ClinicalTrials.gov Identifier: NCT00498368) Rationale Rituximab (Rituxan®, MabThera® and Zytux®), a chimeric monoclonal antibody directed against the CD20 antigen of B cells, causes B‐cell depletion and has proven efficacy in treating B cell lymphomas, leukaemias, transplant rejection and autoimmune disorders including SLE, rheumatoid arthritis, dermatomyositis and ANCA‐associated vasculitis.54 B cell depletion is clearly an attractive therapeutic approach in IgAN as this should switch off production of poorly galactosylated IgA1 and glycan‐specific IgG/IgA autoantibodies and limit the formation of circulating immune complexes. There have, however, been few studies of the use of rituximab in IgAN. In a case series reported by Sugiura et al., the effects of a single‐dose of rituximab (375 mg/m2) was evaluated in 24 patients with primary glomerular diseases, of which five patients had IgAN.55 In this small observational study, there was no benefit from rituximab over conventional treatment in patients with IgAN with regard to proteinuria reduction at 6 month follow‐up, despite a significant reduction in B‐cell numbers. Trial design, inclusion and exclusion criteria, and primary end point A phase IV multi‐centre, randomized, prospective, open‐label trial for the treatment of progressive IgAN with rituximab is currently underway in the United States, with a recruitment target of 54 patients (Table 6). Similar to the SIGN study, patients will undergo a repeat renal biopsy after treatment to examine the effects of rituximab on histopathology. Trial progress to date The trial started in February 2009 and has completed recruitment. The study is expected to report in late 2015. Bortezomib in IgAN (ClinicalTrials.gov Identifier: NCT01103778) Rationale In normal cells, proteasomes regulate protein expression and function by degradation of ubiquitylated proteins. A specific form of proteasome, the immunoproteasome, found exclusively in antigen presenting cells (APC), is capable of generating peptides which are of optimal size and composition for MHC binding. Expression of the immunoproteasome is induced by IFN gamma during an immune response, and excessive immunoproteasomal activity has been linked to inflammatory and autoimmune diseases including SLE and rheumatoid arthritis. Increased immunoproteasome activity in APC results in upregulation of the activated form of NF‐κB, an anti‐apoptotic and pro‐inflammatory regulator of cytokine expression. There is also an emerging evidence for a role of increased immunoproteasome activity in IgAN.56 Excessive switching from proteasome to immunoproteasome in peripheral blood mononuclear cells has been observed in patients with IgAN with high levels of proteinuria.57 The trigger for this switch is not clear, but it has been postulated that an aberrant response to mucosal immune challenge, such as to viral infections that trigger IFN gamma release, may be involved. Bortezomib (Velcade®) is the most widely used proteasome inhibitor in current clinical practice and is licensed for the treatment of multiple myeloma and relapsed mantle cell lymphoma. Multiple mechanisms are likely to be involved in the B cell/plasma cell killing, but ultimately, it is thought bortezomib prevents degradation of pro‐apoptotic factors, permitting activation of programmed cell death in targeted cells. The selectivity of bortezomib for B lineage cells makes it an attractive proposition as a novel treatment in IgAN, although the side effect profile, which includes peripheral neuropathy, myelosuppression causing neutropenia and thrombocytopenia and gastrointestinal effects, may limit its utility to only those patients at greatest risk of progressive renal failure. Trial design, inclusion and exclusion criteria, and primary end point A single‐centre, open‐label, exploratory study examining the effects of bortezomib in IgAN is currently underway in the United States. The details of the study are listed in Table 7. Trial progress to date Recruitment commenced in July 2010 and is currently ongoing; the study is expected to report in late 2016. Potential New Pathways With an increasing understanding of mucosal immunology and the progressive translation of these findings to IgAN, potential new therapeutic targets are continually emerging. At the same time, investigators are looking at whether it might be possible to use both existing and novel therapies developed for other autoimmune diseases or those targeting B cell malignancies to treat IgAN. Manipulation of toll‐like receptor activation in IgAN Toll‐like receptors (TLRs) play a critical role in the early innate immune response to invading pathogens by sensing microbial pathogens and endogenous danger signals via recognition of a diverse range of pathogen‐associated molecular patterns (PAMPs), such as bacterial lipopolysaccharide, RNAs, and DNAs and danger‐associated molecular patterns (DAMPs). TLRs are found on a diverse range of cells including macrophages and dendritic cells, and stimulation of TLRs by the corresponding PAMPs or DAMPs initiates signalling cascades that result in a variety of cellular responses including the production of interferons (IFNs), pro‐inflammatory cytokines and effector cytokines that direct the adaptive immune response. There is increasing interest in the role of TLRs in both maintenance of normal mucosal immunity and the aberrant mucosal responses observed in IgAN. In a murine model of IgAN, B cell expression of TLR‐9 and the signalling molecule, myeloid differentiation factor 88 (MyD88) was significantly greater in mice that were conventionally housed (and therefore exposed to environmental antigens) than in mice housed in pathogen‐free conditions.58 This increased expression of TLR‐9 and MyD88 was associated with more severe IgA‐mediated renal injury. Furthermore, nasal challenge with TLR‐9 ligands aggravated the renal injury and resulted in strong Th1 polarization and increases in serum and mucosal IgA levels. An association between polymorphisms in the TLR‐9 gene and disease progression in two cohorts of patients with IgAN has also been reported. Importantly, TLR‐9 is expressed on mucosal B cells, and it has been suggested that B cell activation through TLR‐9 may be pivotal in the generation of the mucosal IgA response and therefore may be implicated in the generation of poorly galactosylated IgA1. TLR‐9 expression by dendritic cells has also been reported, and activation of dendritic cells through TLR‐9 ligation has been proposed as a potent stimulator of autoantibody production in IgAN.59 Targeting B‐cell TLR activation, therefore, may offer an alternative strategy for modulating mucosal B‐cell activation, poorly galactosylated IgA production and IgA immune complex formation in IgAN. In SLE, it has been shown that signalling through TLR‐7 and TLR‐9 directly antagonizes the immunosuppressive effect of corticosteroids. Furthermore, dual TLR‐7 and TLR‐9 inhibitors alleviate this antagonism and allow lower doses of corticosteroids to be used in SLE without loss of efficacy.60 Hydroxychloroquine (Plaquenil®), a drug commonly used in SLE, is a potent inhibitor of TLR‐9 and, to a lesser extent, TLR‐7 and TLR‐8. In addition, hydroxychloroquine interferes with functioning of the immunoproteasome and antigen presentation by APC. While not a new drug, perhaps it is time to consider a trial of hydroxychloroquine, either alone, or in combination with Nefecon®, to specifically target the mucosal IgA immune system in IgAN. Dissolution of mesangial IgA deposits with IgA1 proteases In order to circumvent the mucosal IgA immune system, several species of pathogenic bacteria including Haemophilus influenzae, Streptococcus pneumonia, Neisseria gonorrhoeae and Neisseria meningitides have evolved the ability to synthesize potent IgA1 proteases that are designed to proteolytically degrade human mucosally secreted IgA1, specifically targeting the O‐glycosylated IgA1 hinge region. These bacterial IgA1 proteases offer an interesting proposition. If it is not possible to completely turn off IgA immune complex formation in IgAN, perhaps it might be possible to use these IgA1 proteases to directly target the IgA1 molecule itself and degrade circulating IgA immune complexes and dissolve away mesangial IgA deposits.61 In a proof‐of‐concept study, Lamm et al. demonstrated that the H. influenzae IgA1 protease could cleave human IgA1 and IgA1‐containing immune complexes in vitro. When systemically administered in a passive mouse model of IgAN, the IgA1 protease significantly reduced the extent of mesangial IgA immune complexes.62 How these observations will translate into clinical trials in humans is currently unclear, but interestingly, in 2012, Shire acquired a worldwide exclusive license from IGAN Biosciences to develop and commercialize protease‐based therapeutics for the treatment of IgAN.61 Conclusion In the relatively short history of clinical trials in IgAN, there has not been a more exciting time. There is worldwide recruitment of IgAN patients to a number of well designed, appropriately sized and powered randomized controlled clinical trials evaluating the effectiveness of both traditional and novel immunomodulatory agents in IgAN. Furthermore, advances in our understanding of the pathogenesis of IgAN are identifying more potential biochemical pathways that might be amenable to therapeutic manipulation. Many of the trials discussed in this review will not report for a number of years, but it is hoped that some of these will provide sufficient evidence to translate into meaningful new treatments for our patients with IgAN in the next 3–5 years. References Notes : BP, blood pressure; GFR, glomerular filtration rate; IgAN, IgA nephropathy. ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; BP, blood pressure; ESRD, end‐stage renal disease; GFR, glomerular filtration rate; IgAN, IgA nephropathy. ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; BP, blood pressure; GFR, glomerular filtration rate; IgAN, IgA nephropathy; UPCR, urine protein : creatinine ratio; UTP, urine total protein. ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; BP, blood pressure; GFR, glomerular filtration rate; IgAN, IgA nephropathy. ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; GFR, glomerular filtration rate; IgAN, IgA nephropathy. ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; EPA, eicosapentaenoic acid; GFR, glomerular filtration rate; IgAN, IgA nephropathy; RAS, renin–angiotensin system. ACEi, angiotensin‐converting enzyme inhibitor; ARB, angiotensinogen receptor blocker; IgAN, IgA nephropathy. Citing Literature Number of times cited: 6 Bengt C Fellström, Jonathan Barratt, Heather Cook, Rosanna Coppo, John Feehally, Johan W de Fijter, Jürgen Floege, Gerd Hetzel, Alan G Jardine, Francesco Locatelli, Bart D Maes, Alex Mercer, Fernanda Ortiz, Manuel Praga, Søren S Sørensen, Vladimir Tesar and Lucia Del Vecchio, Targeted-release budesonide versus placebo in patients with IgA nephropathy (NEFIGAN): a double-blind, randomised, placebo-controlled phase 2b trial, The Lancet, 389, 10084, (2117), (2017). Crossref Robert J Wyatt, Are we ready for targeted therapy for IgA nephropathy?, The Lancet, 389, 10084, (2083), (2017). Crossref See Cheng Yeo, Chee Kay Cheung and Jonathan Barratt, New insights into the pathogenesis of IgA nephropathy, Pediatric Nephrology, (2017). Crossref Magdalena Krochmal, Katryna Cisek, Szymon Filip, Katerina Markoska, Clare Orange, Jerome Zoidakis, Chara Gakiopoulou, Goce Spasovski, Harald Mischak, Christian Delles, Antonia Vlahou and Joachim Jankowski, Identification of novel molecular signatures of IgA nephropathy through an integrative -omics analysis, Scientific Reports, 10.1038/s41598-017-09393-w, 7, 1, (2017). Crossref Bing Du, Ye Jia, Wenhua Zhou, Xu Min, Lining Miao and Wenpeng Cui, Efficacy and safety of mycophenolate mofetil in patients with IgA nephropathy: an update meta-analysis, BMC Nephrology, 18, 1, (2017). Crossref Sigrid Lundberg, Emelie Westergren, Jessica Smolander and Annette Bruchfeld, B cell–depleting therapy with rituximab or ofatumumab in immunoglobulin A nephropathy or vasculitis with nephritis, Clinical Kidney Journal, 10.1093/ckj/sfw106, (sfw106), (2016). Crossref